Many current cancer immunotherapies, including adoptive cell transfer, vaccination, and chimeric antigen T cell receptors, require a priori knowledge of the tumor antigen to target. These therapeutics activate a T cell population against one or more chosen antigens uniquely expressed or upregulated by the patient?s tumor. However patient-to-patient variability and the highly polymorphic nature of the major histocompatibility complex limit the widespread use of these personalized approaches. On the other hand, checkpoint blockade therapies block systemically expressed immunosuppressive pathways and therefore not patient-specific. While checkpoint blockade against the programmed death 1 and CTLA-4 antigens have been FDA approved and are extremely successful in a fraction of patients, complete response rates have been as low as 5%, dependent on the cancer, and demonstrate the need for improvement. The goal of the proposed project is to develop a novel particle-based platform, termed immunoswitch, which links checkpoint blockade with the delivery of a T cell co-stimulatory signal. Prototypic immunoswitch nanoparticles have antibodies against tumor-expressed checkpoint molecules (e.g. PD-L1) and T cell expressed co-stimulatory molecules (e.g. 4-1BB) coupled to their surface and stimulate tumor-specific T cells, in vitro and in vivo. Importantly, immunoswitch particles have activity in vivo in multiple different tumor models. The goals of this revised R21 are to further develop this novel approach and the project will progress in two phases. First will be the synthesis with in vitro characterization and optimization of the immunoswitch particle size efficacy in vivo. Because the T cell is receiving a signal 1 and signal 2, which are known to co-localize during stimulation, from two separate entities (i.e. the tumor cell and immunoswitch particle, respectively), particle size will be investigated and optimized in vivo in the first stages of the project. In phase two, immunoswitch particles will be studied mechanistically in vivo. Will address if the effects intratumoral of immunoswitch lead to systemic effects and memory. The mechanism of action of immunoswitch particles will be studied through analysis of effector cell function and phenotype as well as the particle and immune-cell trafficking within the tumor microenvironment during treatment. If successful, the research will result in the engineering of an innovative platform for cancer immunotherapy.